The Epidemiology Branch is conducting a number of birth defect studies in collaboration with the Health Research Board and Trinity College, Dublin, Ireland. The main objective of these studies is to determine the relationship between folate and birth defects. The birth defects studied to date are neural tube defects (NTDs), oral clefts, congenital heart defects, Down syndrome and omphalocele. These studies focus on biochemical factors in the area of folate metabolism, and on genetic mutations in folate related genes associated with birth defects. In the past we have shown that elevated homocysteine is a risk factor for NTDs, that a mutation in the methylenetetrahydrofolate reductase (MTHFR) gene 677C->T is a risk factor for NTDs, and that a small dose of folic acid (100-200 micrograms) can raise red cell folate to levels that can prevent a fifth to almost a half of NTDs. We have shown that methylenetetrahydrofolate reductase (MTHFD), an important gene in the production of purine and pyrimidine for DNA synthesis, is a risk factor for NTDs. We have shown that auto-antibodies to the folate receptor, previously considered to be an important risk factor for NTDs, are no more common in affected pregnancies than unaffected pregnancies. We have reported that women with vitamin B12 levels in the deficient or borderline range are significantly more likely to bear childrene with NTDs. We have recently expanded our investigation of the possible relationship between maternal folate status and birth defect risks by measuring plasma folate and vitamin B12 levels in women carrying a fetus with a non-neural tube defect malformation. Maternal folate and B12 levels were not significantly lower in mothers carrying affected fetuses for any of the birth defects (other than neural tube defects). Surprisingly, B12 levels were significantly higher in mothers of two case groups: cleft palate and musculoskeletal defects. This may be a chance finding but requires additional investigation. To identify genetic risk factors for NTDs, we evaluated potentially functional single nucleotide polymorphisms (SNPs) that are biologically plausible risk factors for NTDs but that have never been investigated for an association with NTDs, examined SNPs that previously showed no association with NTDs in published studies, and tried to confirm previously reported associations in folate-related and non-folate-related genes. We investigated 64 SNPs in 34 genes for association with spina bifida in up to 558 case families (520 cases, 507 mothers, 457 fathers) and 994 controls in Ireland. Case-control and mother-control comparisons of genotype frequencies, tests of transmission disequilibrium, and log-linear regression models were used to calculate effect estimates. Spina bifida was associated with over-transmission of the LEPR (leptin receptor) rs1805134 minor C allele and the COMT (catechol-O-methyltransferase) rs737865 major T allele Consistent with previous reports, spina bifida was associated with MTHFR 677C>T, T (Brachyury) rs3127334, LEPR K109R, and PDGFRA promoter haplotype combinations. The associations between LEPR SNPs and spina bifida suggest a possible mechanism for the finding that obesity is a NTD risk factor. The association between a variant in COMT and spina bifida implicates methylation and epigenetics as possible mechanisms. Our genome wide association study data enabled us to address an important public health issue regarding food fortification with folic acid. Several studies of elderly individuals found that those who had high blood folate levels in conjunction with low vitamin B12 status had more severe B12 problems including anemia and alterations of B12 metabolism. In our cross-sectional study, 2507 university students provided data on medical history and exposure to folic acid and vitamin B-12 supplements. Blood was collected to measure serum and red blood cell folate (RCF), hemoglobiin, plasma homocysteine (tHcy), methylmalonic acid (MMA), holootranscobalamin, and ferritin in serum. In subjects with low vitamin B-12 concentrations (<148 pmol/L), those who had high folate concentrations (>30 nmol/L;group 1) did not show greater abnormalities in vitamin B-12 cellular function in any area than did those with lower folate concentrations (&#8804;30 nmol/L;group 2). Group 1 had significantly higher holotranscobalamin and RCF, significantly lower tHcy, and nonsignificantly lower (P = 0.057) MMA concentrations than did group 2. The groups did not differ significantly in hemoglobin or ferritin. Compared with group 2, group 1 had significantly higher mean intakes of folic acid and vitamin B-12 from supplements and fortified food. In this young adult population, high folate concentrations did not exacerbate the biochemical abnormalities related to vitamin B-12 deficiency. These results provide reassurance that folic acid in fortified foods and supplements does not interfere with vitamin B-12 metabolism at the cellular level in a healthy population.